Immunoprotection Test Of Staphylococcus Aureus LukF And LukS

Staphylococcus aureus (S.aureus) is an important pathogen that causes diseases among human and animals. Extracellular toxins of S. aureus are the main virulent factors in which leukocidin can induce lysis of the leucocyte by forming a transmembrane pore. Leukocidin consists of two protein components, LukF and LukS, which assemble into a complex on leukocyte membrane in a definite ratio forming a transmembrane pore. But the leukocidin's pathogenesis and, as an immunogen, its immunoprotection were rarely researched.In this experiment, the expressed LukF and LukS proteins were purified and were prepared separately as immunogens for rabbit immunization. Healthy rabbits were divided into LukF protein immuno-group, LukS protein immuno-group, LukF plus LukS protein immuno-group and placebo group. Each group of 6 rabbits were immuned in subcutaneous with 0.5mg immunogen. They were boosted after initial immunity at 3-week interval. Each rabbit was challenged with S.aureus Wood46 at 5×107.1CFU 3-week later after secondary immunization. The clinical signs and body temperature of the rabbits were observed and recorded for the next 5 days. Then the rabbits were killed and Pathological lesions of organs were recorded. The microscopic pathology of the organs with lesions was detected. Serum samples from each rabbit were taken just before the inoculation and at 1-week intervals after that for detecting antibody titer. The results were as follows: Luk as an immunogen could provide to a certian extent immunoprotection from tissue necrosis in some organs, such as kidney. Immunogens LukF and LukS respectively provided 3/6 rabbits immunoprotection, and Immunogen LukF plus LukS provided 5/6 rabbits immunoprotection, and placebo provided 0/6 rabbits immunoprotection. At the third week after secondary inoculation, Sera antibody titers of rabbits from LukF plus LukS immuno-group were 1:64 000 to 1: 256 000, and that of rabbits both from LukF and LukS immuno-groups was 1:16 000 to 1:64 000.To research on the immunoprotection mechanism of leukocidin, the mixing proportion of the purified LukF and LukS proteins for perforation on cell membrance was firstly studied. The optimal mixing proportion of that was confirmed to be 1:1. Then perforation forming and leucocyte destruction of LukF and LukS could be prevented by the specific antibodies to LukF and LukS in vitro. On the basis of results above, the rabbits immuned with LukF and LukS protein individually and both, were challenged with S.aureus Wood46, and the blood was collected on fifth days after challenge for detecting leucocyte numbers. the leucocyte numbers from healthy rabbits were (5.87±0.24)×10~6/L, that from rabbits inoculated with LukF protein was (7.24±0.47)×10~6/L, that from rabbits inoculated with LukS protein was (7.2±0.30)×10~6/L, and that from rabbits inoculated with LukF plus LukS was (8.52±0.52)×10~6/L. It was demostrated that the Luk antibody could effectively protect leucocyte from destruction by leukocidin in vivo.All above results illustrated that S.aureus LukF and LukS protein has favorable immunogenicity, which can cause favorable immune response and protect leucocyte from destruction and tissue necrosis effectively.